Scientists use `reverse genetics' to develop bird flu vaccine

WASHINGTON—Scientists who are racing to create vaccines to protect humans from the vicious bird-flu virus that's sweeping across Asia and Eastern Europe are using an ingenious strategy known as "reverse genetics."

The idea is to take apart the genes that make up two viruses—one deadly, the other relatively harmless—and reassemble parts of them into a new, weakened virus that can be used safely as a vaccine.

The government already has ordered 20 million doses of such a recombined vaccine, even though it's still being tested and hasn't been approved for human use. President Bush is scheduled to announce his plan to meet the bird-flu threat Tuesday.

Reverse genetics is "the only efficient way to produce vaccine strains" for the deadly bird-flu virus, said Yoshihiro Kawaoka, a viral expert at the University of Wisconsin, Madison.

As of Oct. 27, 121 people in Southeast Asia had contracted flu from chickens or other birds and 62 had died, a mortality rate of 51 percent, according to the World Health Organization.

So far, there've been only two confirmed cases, both in Thailand, in which the virus spread from one person to another. Both involved close relatives of people who had the disease.

But scientists fear the virus might mutate at any time into a form that could pass easily between humans. That's what happened in the terrible Spanish Flu of 1918, when as many as 50 million people perished worldwide.

A vaccine is available against the ordinary, seasonal flu that strikes each winter, killing an estimated 36,000 people in the United States alone. But as yet there's no vaccine that's proved to be effective for humans against the lethal new bird-flu strain, H5N1.

The virus' genes are made of RNA, a simpler form of DNA, the genetic blueprint for all complex organisms. There are eight RNA genes in the H5N1 virus, but only two of them cause infection.

The symbol H5 refers to the fifth variant of a gene that produces a protein, called hemagglutinin (pronounced he-mah-GLUE-tin-in), which sits on the surface of a virus and enables it to enter and infect a healthy cell. The N1 gene makes another protein, neuraminidase (new-rah-MINI-dace), which allows new viruses to escape from an infected cell and infect others.

A candidate H5N1 vaccine developed at St. Jude Children's Research Center in Memphis, Tenn., has passed initial safety tests and is being tried out in a small number of humans. The National Institute of Allergy and Infectious Diseases, a government agency in Bethesda, Md., is working on vaccines for other variants that may develop in the future.

One problem with these vaccines is that they're designed to protect against the form of the virus that infects birds, and they may not work well against a different, human variety. If so, scientists will have to create a new vaccine and start production over again.

Here's how the reverse-genetic process works, as explained by Richard Webby, a leading vaccine researcher at the St. Jude laboratory:

_ The first step is to obtain a sample of the H5N1 virus from the World Health Organization, extract its RNA and reproduce it.

_ Next, researchers take six genes from the common, seasonal virus strain known as H1N1, which is now circulating in the world. These genes perform basic housekeeping functions in the virus, and aren't dangerous by themselves.

_ Finally, the N1 gene is added to the mix, creating a reconstituted H5N1 virus. Quantities of the virus can be grown in eggs or in cells from a dog or monkey kidney. Other substances are added to complete the vaccine.

When the vaccine containing the weakened virus is injected into an animal or person, it tricks the body into thinking it's been infected. The body's natural immune system detects the H5 protein and churns out vast numbers of protective particles called antibodies. The antibodies latch onto the viruses and prevent them from entering a target cell.

Once a vaccine has been designed and tested, it needs to be produced in sufficient quantities. Only nine companies in the world—three of them in the United States—are capable of large-scale vaccine production, according to Dr. Tara O'Toole, the director of the Center for Biosecurity at the University of Pittsburgh Medical Center.

"We could make the first dose of vaccine in six weeks, and tens of millions of doses in three months after that," O'Toole said. "It takes months to make a vaccine."

For this year, it's already too late, O'Toole said.

A further complication is the fact that at least two doses of vaccine will be required, each six times more powerful than the common vaccine that's used against seasonal flu, according to Michael Osterholm, a bird-flu expert at the University of Minnesota in Minneapolis. That puts an even greater burden on already overstretched vaccine manufacturers.

"We will have enough vaccine in about a year for 1 billion doses," Osterholm said. That would be enough to cover 500 million people, less than 10 percent of the world's population.

"The good news is we can make a vaccine," Osterholm said. "The bad news, it's not going to prevent a pandemic in the next 12 to 18 months."

Antiviral drugs such as Tamiflu and Relenza are also in short supply. They're used principally to mitigate the effects of flu, but may also help prevent infection.

The standard way to produce flu vaccine is by growing the weakened virus in chicken eggs. This takes at least six months. An alternative method is to use mammal cells, usually taken from monkey kidneys.

British scientists are testing another type of flu vaccine that consists of DNA rather than RNA. They say it could be produced in large vats owned by pharmaceutical manufacturers in the United States and Europe in two or three weeks instead of months.

"In an influenza pandemic, a DNA vaccine would not need to be cheaper or better than a conventional (virus-based) vaccine, but it must be capable of fast production," University of London biochemist Peter Dunnill wrote in the November-December issue of the journal Biotechnology Progress.

To make a DNA vaccine, scientists insert the instructions for making the H5 protein into a small ring of DNA taken from the common intestinal bacteria E. coli. When a vaccine containing that bit of DNA is injected into a cell, it triggers the immune system much as a conventional vaccine does.

"At present almost all of the focus of pandemic planning is on virus-based vaccines," Dunnill said. "A DNA vaccine is not a panacea. However, it could be useful if the situation gets out of hand."